Cytosolic Alkalinization of Vascular Endothelial Cells Produced by an Abrupt Reduction in Fluid Shear Stress

From the Department of Medicine (R.C.Z., M.C.C.), Division of Cardiology, Johns Hopkins Bayview Medical Center, Johns Hopkins University School of Medicine, Baltimore, Md; the Laboratory of Cardiovascular Science (R.C.Z., P.S.B., L.C., M.C.C.), Gerontology Research Center, National Institute on Aging, National Institutes of Health, Baltimore, Md; and the Laboratorio di Patologia Vascolare (M.C.C.), Istituto Dermopatico dell' Immacolata, Rome, Italy.

Correspondence to Roy C. Ziegelstein, MD, Department of Medicine, Division of Cardiology, Johns Hopkins Bayview Medical Center, 4940 Eastern Ave, Baltimore, MD 21224-2780. E-mail rziegels{at}welchlink.welch.jhu.edu

Abstract—Reductions in fluid shear stress produce endothelium-dependent vasoconstriction and promote neointimal hyperplasia, but the intracellular signaling mechanisms involved in these processes are poorly understood. To examine whether decreases in fluid shear stress affect endothelial cytosolic pH, carboxy-seminaphthorhodafluor-1–loaded rat aortic endothelial cells were cultured in glass microcapillary tubes and examined during abrupt reductions in laminar flow. After a 30-minute exposure to a shear stress of 2.7 dyne/cm² in bicarbonate buffer, the acute reduction of fluid shear stress from 2.7 to 0.3 dyne/cm² transiently increased cytosolic pH from 7.20±0.02 to 7.47±0.07 (mean±SEM, P<.05 versus control). This was not affected by prior inhibition of the Na⁺-H⁺ exchanger with 10 µmol/L ethylisopropylamiloride but was abolished in bicarbonate-free buffer. Recovery from an ammonium chloride prepulse–induced acid load occurred more rapidly when fluid shear stress was abruptly reduced from 2.7 to 0.3 dyne/cm² after maximal acidification (+0.04±0.02 pH unit at 2 minutes) than when shear stress was maintained at 2.7 dyne/cm² continuously (0.00±0.00 pH unit at 2 minutes, P<.05). This accelerated cytosolic pH recovery was dependent on the presence of bicarbonate ion and was blocked by the addition of the exchange inhibitors DIDS (100 µmol/L) and ethylisopropylamiloride or by removal of buffer Na⁺, indicating that the acute reduction in fluid shear stress activates the extracellular Na⁺–dependent Cl^--HCO₃^- exchanger and the Na⁺-H⁺ exchanger and increases cytosolic pH in vascular endothelial cells.

Key Words: endothelium • cytosolic pH • shear stress • carboxy-seminaphthorhodafluor-1 • DIDS

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